There have been attempts at in vivo apoptosis detection and imaging using Annexin V and Annexin V derivatives (see, e.g., Kietselaer et al., 2003; Belhocine et al., 2004; Reddy, 2005; Boersma et al., 2005; Watanabe et al., 2006; Vanderheyden et al., 2006; and Corsten et al., 2006). Other attempts using novel compounds that rely on the perturbation and alterations of the normal organization of the cell plasma membrane have also been attempted (see, e.g., U.S. Patent Application Publications 2005/0244812 and 2005/0276750).
None of the previously described methods uses a cell permeant probe for the detection and imaging of apoptosis. As a result of this and other issues, all of the aforementioned methods have been plagued with problems resulting in high backgrounds and lack of binding to certain apoptotic tumor cells. Annexin V is not cell permeant, is slow to penetrate any tissues, has high background, and does not detect early apoptotic cells (Kietselaer et al, 2003; Belhocine et al., 2004; Boersma et al., 2005; Watanabe et al., 2006; Vanderheyden et al., 2006, and Corsten et al, 2006). Leading to high background, Annexin V binds positively to normal and healthy bone marrow derived cells (Dillon, 2001). It has been reported that Annexin V does not bind to all tumor cells (Dicker, 2005).
In the Ziv publications (U.S. Patent Application Publications 2005/0244812 and 2005/0276750), it is reported that their compounds accumulate in apoptotic cells at a rate faster than they accumulate in cells that are not undergoing cell wall turnover. This leads to high background levels and lack of specificity similar to Annexin V. Requiring a compromised cell state also prohibits the detection of cells that are in the early stages of apoptosis.
Thus, previously described methods of in vivo apoptosis detection and imaging lack specificity and sensitivity and are subject to high background. The use of sensitive and specific cell permeant inhibitor probes that bind to specific active enzymes and proteases involved in apoptosis has not been described.
Thus, methods and products for in vivo determination of the apoptotic state of cells in an organism are needed.